Molecular chaperons are proteins which mediate protein folding. They bind non-covalently to exposed surfaces of proteins that are newly synthesized or are denatured or misfolded, and assist them to fold into correct conformations. Molecular chaperons are also involved in a number of cellular processes such as protein synthesis, protein translocation and DNA replication.
Molecular chaperons include heat shock proteins, which are proteins whose expression increases significantly in cells following an exposure to unusually high temperature (heat shock) or an exposure to a wide variety of physiological stresses. This increase in the molecular chaperon expression in turn provides cells with protection against the adverse effects of hyperthermia, as demonstrated by the thermotolerance of cells for otherwise lethal temperatures if the cells are pre-conditioned by a brief exposure to high temperature.
Physiological stresses inducing heat shock protein expression include a wide variety of pathological conditions associated with many diseases. The synthesis of heat shock proteins in cells exposed to such stresses, indicates the protection of the cell against the physiological stresses, like also in the case of the heat shock response.
One such pathological condition associated with induction of molecular chaperons is ischemic injury. Ischemic injury to tissues results from deterioration of blood supply for any possible. For instance, prolonged coronary occlusion causes severe damage to myocardium, leading to myocardial necrosis and jeopardizing the chances for recovery even if the blood flow is restored. In brain, to significant damages may frequently be caused by ischemia, leading to death of the brain-tissue.
It was observed that the amount of heat shock protein hsp70 increased in the myocardium during ischemia leading to necrosis even if the duration of ischemia is short. In these cases, likewise in a heat shock, the enhanced hsp70 content of the cells protects the same against the consequences of a next ischemia, which would otherwise cause necrosis (DAS, D. K., et al. Cardiovascular Res.: 578, 1993). It has also been observed when rat cells in culture were subjected to ischemia, J. Clin. Invest., 93: 759–767 (1994)). Accordingly, heat shock proteins synthesized by myocardial cells provide protection against ischemic injuries.
The situation in brain-tissue is similar, wherein cerebral ischemia results in increased expression of heat shock protein in the brain tissue. Experiments have also proved that pretreatment of animals with sub-lethal ischemia induces heat stress protein (hsp70) and protects the brain against more severe subsequent ischemic insult. (Simon, et al., Neurosci. Lett., 163:135–137 (1993)).
Yet another example of physiological stress on tissues and organs associated with molecular chaperon induction is provided by inflammatory diseases. Inflammation is a non-specific response of host cells to entry of foreign material such as in case of infection by various bacterial and viral pathogens, and involves aggregation and activation of leukocytes to the injury site, which results in production and release of high levels of reactive oxygen species and cytokines. These cytokines and reactive oxygen radicals attack the pathogen, but also damage the host tissues (Jaquier.Sarlin, Experientia, 50: 1031–1038/1994/). It is believed that as a protection against these toxic mediators of inflammation, the host tissues increase production of molecular chaperons. Molecular chaperons thus produced protect host cells from damages caused by reactive oxygen species and protect cells from cytotoxicity of TNF and other cytokines and reactive oxygen radicals. In animal studies, it has been demonstrated the pre-exposure of an animal to heat shock, with resulting increase of a heat shock protein (hsp70) expression, resulted in remarkable decrease in pulmonary inflammation. Accordingly, molecular chaperons serve anti-inflammatory function.
The above examples illustrate ability of molecular chaperons to protect cells against various physiological stresses disturbing cellular homeostatic balance and causing injury to cells. Molecular chaperons have also been shown to be advantageous in treating neoplasms. For example, it has been reported that when tumor cells are transfixed with a gene encoding a molecular chaperon (65 kd hsp), they lose or show decrease in their tumorigenicity (PCT Application No. PCT/GB93/02339). Furthermore, it has also been reported that tumor cells, in response to heat stress, express molecular chaperons in increased amount. However, they are present not in cyto-plasm, but on the surface of cell membranes. (Ferrarini,M. et al Int.J.Cancer, 51:613–619/1992/). Increased presence of molecular chaperones on cell surfaces correlates with increased sensitivity of NK (natural killer) cells toward the tumor cells, allowing better targeting, infiltrating, and killing of the tumor cells by NK cells (Kurosawa.S. et al. Eur.J.Immunol. 23:1029/1993/).
In view of the advantages associated with increased molecular chaperon expression in cells, a method which increased such expression or increased activity of molecular chaperons would be highly desirable.